Thermodynamic Analysis of the Interaction of Lipopolysaccharides with Cationic Compounds

Abstract

AbstractBacterial lipopolysaccharides (LPS) from Gram‐negative bacteria, located in the outer leaflet of their outer membrane, are called endotoxins due to their ability to induce a variety of biological effects in mammals. Their lipid moiety, lipid A, is called “the endotoxic principle” and is responsible for the toxic effects of LPS. As a result of the polyanionic character of LPS, the study of the interaction with divalent cations Mg2+ and Ca2+ and polycationic peptides such as polymyxin B (PMB) and its nonapeptide PMBN is of considerable interest, and therefore the authors have investigated the interaction of LPS/lipid A with cationic compounds by applying isothermal titration calorimetry (ITC) and differential scanning calorimetry (DSC). The data indicate a clear binding of the divalent cations with the anionic glycolipids, leading to calorimetric reactions, such as an increase in the phase transition temperature, Tm, of the gel to the liquid crystalline phase of LPS, indicating a stabilization of the gel phase. The peptides react quite differently as assessed by DSC. In contrast to the interaction of divalent cations with the glycolipids, a destabilization of the gel phase is observed, accompanied by a decrease in the gel to liquid crystalline phase transition enthalpy for the peptide‐glycolipid interaction. The extent of this effect is peptide‐concentration‐dependent. Using ITC for the analysis of the binding reaction of the cations and the peptides with the glycolipid in the liquid crystalline phase, strong exothermic effects are observed. These are indicative of the dominance of electrostatic attractions between the reaction partners. Interestingly, Ca2+ binding to LPS leads to a slightly exothermic reaction, whereas Mg2+ binding leads to an endothermic reaction (some kJ/mol). The observed highly endothermic binding reactions for the lipid‐peptide interaction in the gel phase are mainly driven by a gain in entropy. This is explained by the fact that during binding, water molecules from the hydration shells of the components are liberated. Although the electrostatic attraction is still the driving force of the interaction, it is quantitatively of minor importance for the interaction in the gel phase. The binding results are discussed in terms of competition between electrostatic interaction and hydration forces. These data are of importance for the understanding of the reaction mechanisms of cationic compounds with LPS under physiological conditions.